Project Summary HIV-infected people whose viral load is below target levels due to effective anti-retroviral therapy (ART) continue to be at increased risk for cardio-pulmonary disease with over 75% of patients with chronic HIV disease showing clinical manifestations. Recent studies in the laboratories of the applicants provided evidence that HIV proteins and in particular HIV-Nef is retained in plasma and lung fluids of HIV patients on effective combined anti-retroviral therapy (ART). Based on this previous work we plan to elucidate in preclinical in vitro and in vivo models the mechanism of endothelia damage and premature aging. Our main hypothesis is that intracellular HIV proteins are released from cells together with HIV-Nef and travel through extracellular vesicles (EV) to cause cardiopulmonary changes leading to comorbidities. In aim 1 we will study HIV-proteins in extracellular vesicles and their association with specific cargo with focus on surface- and intra-vesicular orientation. The detection of surface markers for specific EV-associated HIV proteins, will allow for future therapeutic and diagnostic applications including antibody-based targeting techniques. In aim 2, we will analyze the role EV-associated HIV proteins in delivering HIV-EV-associated cargo throughout the body to increase inflammation and cell senescence. Specifically, we will use multi-antibody panels to determine cell identity and location of the ?homed? EV. In aim 3, we will focus on the pathology of the delivered HIV-EV associated cargo. As a proof of principal we will test specific intervention strategies including ADAM17 inhibitors and senolytic agents in preclinical mouse models for HIV-protein delivery through EV.